A clostridial glycine reductase complex catalyzes the reductive deamination of glycine to acetate and ammonia with the concomitant esterification of orthophosphate and synthesis of ATP. Use of hydrophobic chromatographic matrices such as phenyl and octyl sepharoses has greatly facilitated the separation of the two membrane-associated proteins (B and C) that are components of the glycine reductase complex. In the separation procedures used earlier, the C protein co-migrated with an iron protein. However, using Iron-55 labeled enzyme preparations it has been shown that the C protein does not contain iron. The C protein preparations currently studied are not inhibited by treatment with radical scavengers such as hydroxylamine or hydroxyurea. Thus, the postulated similarity between C protein and E. coli ribonucleotide reductase protein B does not appear to exist. The latter protein contains a tyrosine radical stabilized by ferrous iron which participates in the catalytic reaction. A collaborative project on purification and identification of a rapidly turning over selenocysteine-containing protein synthesized by liver and exported to the serum is in progress with Dr. Raymond Burk of the University of Texas, San Antonio. The biological mechanism of formation of selenocysteine residues in selenium-dependent enzymes is investigated in two different bacterial systems. The selenoprotein component of Clostridium sticklandii glycine reductase has been partially sequenced around the single selenocysteine residue. Synthetic RNA probes corresponding to this amino acid sequence then can be used to isolate the complementary gene sequence which should specify the nature of the amino acid precursor of the selenocysteine residue. A collaborative project with Dr. August Bock of Munchen has a similar objective. Using his cloned gene from E. coli that specifies a selenocysteine-containing subunit of formate dehydrogenase we will provide the requisite amino acid sequence for comparison with the DNA sequence (Bock's part of the project). Studies on the mechanism of methane production from acetate (Gas Research Institute-supported project) have started with isolation and study of carbon monoxide dehydrogenase (a nickel enzyme presumed to be involved).